Christopher T. Rossi

DICER1 mutations in embryonal rhabdomyosarcomas from children with and without familial PPB-tumor predisposition syndrome.

Embryonal rhabdomyosarcoma (ERMS) is the most common childhood sarcoma and is a component of the familial pleuropulmonary blastoma (PPB)‐predisposition syndrome. Using the PPB model, we hypothesized that DICER1 mutations would be found in familial and sporadic forms of ERMS. Blood samples from four children with familial PPB and ERMS, and 52 sporadic ERMS tumors were tested for DICER1 mutations. Germline DICER1 mutations were found in all four patients with familial PPB and 2 of 52 (3.8%) sporadic ERMS had somatic mutations. Our findings confirm the pathogenetic relationship between ERMS and PPB suggesting that ERMS may result from abnormal miRNA regulation. Pediatr Blood Cancer 2012;59:558–560.

Rhabdomyosarcoma of the head and neck in children: review and update.

OBJECTIVE To review the clinical presentation, histology, staging, treatment modalities, and survival for pediatric head and neck rhabdomyosarcoma (non-orbital). STUDY DESIGN Retrospective chart review at a tertiary pediatric hospital of children treated over 18 years (1996-2014) for primary head and neck non-orbital rhabdomyosarcoma. METHODS Medical charts were examined for clinical presentation, staging, histology, genetic abnormalities, treatment modalities, recurrence and complications from treatment. RESULTS Our cohort was 17 children (7 male, 10 female) with rhabdomyosarcoma with a median age of 6.3 years (range <1-19). The majority of tumors were of parameningeal location (13/17) with embryonal histology (11/17). Twenty-nine percent (5/17) demonstrated advanced metastatic disease at initial referral. Fifty-three percent (9/17) had skull base erosion and/or cranial nerve deficits. PET CT scan was performed in 4 patients. The overall survival was 75% for the duration of the study. Primary surgical excision was performed in all 4 patients with nonparameningeal tumors as compared to only 1 patient with a parameningeal tumor. All received chemotherapy and radiotherapy, as none had completely resectable disease. CONCLUSION Pediatric non-orbital primary rhabdomyosarcoma of the head and neck usually has a rapid onset and presents with advanced disease. Our analysis found that the majority of patients in our series had a cranial neuropathy at presentation, which highlights how advanced the disease is in these patients at presentation. The first mode of surgical intervention should be directed toward biopsy in junction with a metastatic work-up. Primary excision with negative microscopic margins for nonparameningeal rhabdomyosarcoma is ideal to spare radiotherapy but was not achievable in our cohort. The benefits of second-look biopsy after chemotherapy and radiation are still unproven; however, we believe that it was beneficial in two patients in our review for further resection thus decreasing subsequent radiation. Fluorodeoxy-d-glucose positron emission tomography (PET) to evaluate disease post treatment may further define the role for second look surgery.

More Than Hypomyelination in Pol-III Disorder

Abstract The 4H syndrome (hypomyelination, hypodontia, hypogonadotropic hypogonadism) is a newly recognized leukodystrophy. The classical form is characterized by the association of hypomyelination, abnormal dentition, and hypogonadotropic hypogonadism, but the recent identification of 2 genes responsible for the syndrome demonstrates that these 3 main characteristics can be variably combined among “Pol-III (polymerase III)–related leukodystrophies.” The pathophysiology of this group of diseases is still to be elucidated, and there are no neuropathologic descriptions of brain tissue. We report the clinical, neuroradiologic, and neuropathologic findings of a patient affected by 4H syndrome with confirmed POLR3A mutations. We found a marked loss of oligodendrocytes, varying in severity in different brain regions, and accompanied by severe loss of myelin, moderately severe loss of axons, and patchy perivascular regions of betterpreserved white matter. There was relatively mild white matter astrogliosis and microgliosis. A macrophage reaction involving viable normal-appearing oligodendroglia suggests the possibility of an immunologic process in this disorder. Cortical laminar astrogliosis and mineralization of Layers I and II in particular were present. Thus, despite theuniformly hypomyelinating pattern seen on magnetic resonance imaging, neuropathologic examination reveals a complex heterogeneous leukodystrophy with prominent neuroaxonal and glial involvement in this disorder.

Solid pseudopapillary tumor in a pancreatic rest of the jejunum

Solid pseudopapillary tumors of the pancreas (SPTP) are very rare, and an SPTP arising in a pancreatic rest has been reported only 4 times previously and never in association with the jejunum. We report this unusual case of a 16 year old girl who presented with 4 days of intermittent, crampy abdominal pain and was found to have an SPTP arising in a pancreatic rest of the jejunum.

DYSREGULATION OF UPSTREAM AND DOWNSTREAM TRANSFORMING GROWTH FACTOR-β TRANSCRIPTS IN LIVERS OF CHILDREN WITH BILIARY ATRESIA AND FIBROGENIC GENE SIGNATURES

INTRODUCTION Our previous work demonstrated that the transforming-growth factor (TGF) β pathway plays a central role in the liver fibrosis associated with experimental biliary atresia (BA). To confirm these findings in humans, we performed an in silico analysis of publicly available microarray data from liver specimens from children with BA, with the hypothesis that the TGF-β pathway would be dysregulated. METHODS We analyzed publicly available liver gene expression microarray data from 47 infants with BA. We re-analyzed the microarray image files and clinical data to compare gene expression differences between the fibrogenic and inflammatory cohorts identified in the initial study. Targets from the microarray analysis were confirmed using the animal model of BA. RESULTS Analysis of variance (ANOVA) detected 6903 transcripts (2822 distinct genes) differentially regulated between groups (p < 0.01; fold change >1.2). We used a targeted approach to identified a subgroup of 24 TGF-β-related transcripts. Expressions for procollagen transcripts were increased in the fibrogenic group (1.2-fold to 1.4-fold); expression of matrix metalloproteinase (MMP)-7 was similarly increased 2-fold, while MMP-9 and plasminogen activator inhibitor-1 were decreased 2-fold and 3-fold respectively. Integrins β5 (1.18-fold) and β8 (1.84-fold) also demonstrated increased expression in the fibrogenic group. Increased expression of β5 (3-fold) and β8 (5-fold) as well as Smad-3 (4-fold) and Smad interacting protein (SIP)-1 (3.5-fold) mRNA was confirmed in experimental BA. Phosphorylated Smad-3 protein in the experimental group was also nearly twice that of the control group, further implicating the TGF-β pathway. CONCLUSION Gene transcripts for known upstream and downstream TGF-β mediators are differentially expressed in liver specimens from children with BA and a fibrogenic gene signature. The same integrins that were dysregulated in the human specimens were also found to be up-regulated in our animal BA model, as were other intermediaries in the TGF-β pathway. Further investigation into whether these mediators may be attractive targets for future therapy in children with BA is warranted.

Parathyroid carcinoma in a child: an unusual case of an ectopically located malignant parathyroid gland with tumor invading the thymus

Parathyroid carcinoma is exceptionally rare in children. Parathyroid carcinoma in the pediatric population most typically presents with significant hypercalcemia and a palpable neck mass. The authors report the seventh case of parathyroid carcinoma diagnosed in a child younger than 16 years. To our knowledge, this case is the first documented case, in the pediatric population, of parathyroid carcinoma in an ectopically located parathyroid gland with tumor invading the thymus.

Integrin β-8, but not β-5 or -6, protein expression is increased in livers of children with biliary atresia

Objectives: Our previous work demonstrated altered messenger RNA expression of integrin &bgr;-5 and -8, using an in silico analysis of publically available data from patients with biliary atresia (BA); however, we were unable to demonstrate statistically significant differences in protein expression because of sample size. In the present study, we repeated the analysis of liver fibrosis and protein expression of the integrins in a larger cohort of patients with BA and compared them with patients undergoing liver biopsy for other diagnoses, with the hypothesis that ≥1 of the integrins would be differentially expressed. Methods: Liver specimens were obtained at 2 collaborating institutions. Samples from infants with BA (n = 23) were compared with samples from those who underwent liver biopsy for neonatal hepatitis (n = 9). All of the specimens were analyzed by 2 pathologists (C.R. and R.A.), who were blinded to the diagnoses. Standard Ishak scoring was performed to evaluate fibrosis and inflammation, and immunohistochemical (IHC) positivity was graded from 0 to 4. Comparisons between the IHC positivity and Ishak scoring for the BA and control groups were performed using the Student t test with P < 0.01 considered significant because of the multiple comparisons. Interobserver variability was assessed by intraclass correlation (ICC). Results: Pooled analysis from specimens from patients with BA showed significantly more fibrosis than controls based on Ishak scores (3.21 ± 1.82 vs 1.17 ± 1.00, P < 0.005). IHC evaluation showed increased integrin &agr;&ngr;&bgr;8 protein expression when compared with controls (2.67 ± 0.81 vs 1.72 ± 0.62, P < 0.005); however, there were no significant differences in integrin &agr;&ngr;&bgr;5 (1.93 ± 0.84 vs 1.50 ± 0.90, P = 0.23) or integrin &agr;&ngr;&bgr;6 (0.85 ± 1.20 vs 0.94 ± 0.85, P = 0.82) expression. These data were confirmed on individual analysis. Interobserver agreement was fair for integrin &agr;&ngr;&bgr;5 (ICC 0.52), good for integrin &agr;&ngr;&bgr;6 (ICC 0.72), and excellent for integrin &agr;&ngr;&bgr;8 (ICC 0.79) and fibrosis (ICC 0.89). Conclusions: Our data show that integrin &agr;&ngr;&bgr;8, but not integrin &agr;&ngr;&bgr;5 or integrin &agr;&ngr;&bgr;6, protein expression is increased in liver specimens of patients with BA. These data support the mounting evidence that transforming growth factor-&bgr; (TGF-&bgr;) activation is responsible for the fibrosis found in BA. Anti-integrin &agr;&ngr;&bgr;8 or more global integrin blocking strategies may be therapeutic options in BA, but further work is clearly needed.

PD-L1 checkpoint inhibition and anti-CTLA-4 whole tumor cell vaccination counter adaptive immune resistance: A mouse neuroblastoma model that mimics human disease

Background Adaptive immune resistance induces an immunosuppressive tumor environment that enables immune evasion. This phenomenon results in tumor escape with progression and metastasis. Programmed cell death-ligand 1 (PD-L1) expressed on tumors is thought to inhibit tumor-infiltrating lymphocytes (TILs) through programmed cell death 1 (PD1), enabling adaptive immune resistance. This study investigates the role of PD-L1 in both mouse and human neuroblastoma immunity. The consequence of PD-L1 inhibition is characterized in the context of an established whole tumor cell vaccine. Methods and findings A mouse model of neuroblastoma was investigated using an Id2 knockdown whole cell vaccine in combination with checkpoint inhibition. We show that immunogenic mouse neuroblastoma acquires adaptive immune resistance by up-regulating PD-L1 expression, whereas PD-L1 is of lesser consequence in nonimmunogenic neuroblastoma tumors. Combining PD-L1 checkpoint inhibition with whole tumor cell/anti-CTLA-4 vaccination enhanced tumor cell killing, cured mice with established tumors, and induced long-term immune memory (6 months). From an evaluation of patient neuroblastoma tumors, we found that the inflammatory environment of the mouse neuroblastoma mimicked human disease in which PD-L1 expression was associated directly with TILs and lower-risk tumors. High-risk patient tumors were lacking both TILs and PD-L1 expression. Although a correlation in immunity seems to exist between the mouse model and human findings, the mouse tumor model is induced and not spontaneously occurring, and furthermore, the number of both mouse and human correlates is limited. Conclusions This study demonstrates the role PD-L1 plays in neuroblastoma’s resistance to immunity and defines the nonredundant effect of combination checkpoint inhibition with vaccine therapy in a mouse model. High-risk, nonimmunogenic human tumors display both diminished PD-L1 expression and adaptive immune resistance. Paradoxically, high-risk tumors may be more responsive to effective vaccine therapy because of their apparent lack of adaptive immune resistance.

Solution blow spun polymer: A novel preclinical surgical sealant for bowel anastomoses

BACKGROUND Solution blow spinning is a technique for depositing polymer fibers with promising potential use as a surgical sealant. This study assessed the feasibility and efficacy of solution blow spun polymer (BSP) for sealing bowel perforations in a mouse model of partial cecal transection. We then evaluated its use for reinforcing a surgical anastomosis in a preclinical piglet model. METHODS Three commercially available surgical sealants (fibrin glue, polyethylene glycol (PEG) hydrogel, and cyanoacrylate) were compared to BSP in the ability to seal partially transected cecum in mice. For anastomosis feasibility testing in a piglet model, piglets were subjected to small bowel transection with sutured anastomosis reinforced with BSP application. Outcome measures included anastomotic burst pressure, anastomotic leak rate, 14-day survival, and complication rate. RESULTS For the mouse model, the survival rates for the sealants were 30% for fibrin glue, 20% for PEG hydrogel, 78% for cyanoacrylate, and 67% for BSP. Three of 9 mice died after BSP administration because of perforation leak, failure to thrive with partial obstruction at the perforation site, and unknown causes. All other mice died of perforation leak. The mean burst pressure at 24h was significantly higher for BSP (81mm Hg) when compared to fibrin glue (6mm Hg, p=0.047) or PEG hydrogel (10mm Hg, p=0.047), and comparable to cyanoacrylate (64mm Hg, p=0.91). For piglets, 4 of 4 animals survived at 14days. Mean burst pressures at time of surgery were 37±5mm Hg for BSP and 11±9mm Hg for suture-only controls (p=0.09). CONCLUSIONS Solution blow spinning may be an effective technique as an adjunct for sealing of gastrointestinal anastomosis. Further preclinical testing is warranted to better understand BSP properties and alternative surgical applications.

Inflammatory myofibroblastic tumor as a second neoplasm after Wilms tumor

We report two survivors of Wilms tumor (WT) who developed inflammatory myofibroblastic tumor (IMT). The first patient had IMT in the mainstem bronchus 1 year after WT therapy that included vincristine, dactinomycin, doxorubicin (VDA) and whole lung radiation therapy (XRT). The second patient had IMT in the cecum 7 years after WT therapy consisting of VDA and abdominal XRT. The unlikely co‐occurrence of WT and IMT suggests a genetic link between these tumors or that IMT can arise as a complication of cancer therapy. IMT should be considered in WT survivors who develop secondary tumors, especially at unusual sites. Pediatr Blood Cancer 2015;62:1075–1077.